I had samples processed over 2 MiSeq runs, where all the amplicons (515F-806R) were pooled into one library and sequenced twice. I intend on merging the data sets for analysis.
Now, I have read through the two threads:
But still, have questions:
So far, my methodology has included creating a manifest to join all the paired end reads.
Due to sequencing each sample on two runs, I decided to provide each its own unique sample ID.
For example, from the first sequencing run, “Site 1” will have Forward and Reverse reads, and the second sequencing run is titled “Site 1b” as its sample ID with its Forward and Reverse reads.
Since I am using only one manifest, all denoised/deblurred samples will be compiled into one output of rep-seq.qza and table.qza files once completed.
Is there a way of combining “Site 1” and “Site 1b” at this time? If so what commands are suggested?
I noticed that the “Fecal microbiota transplant” tutorial has the following code:
qiime feature-table merge
qiime feature-table merge-seqs
However, my interpretation of its usage from the two mentioned forum discussions above would be:
You have to make two manifests, where each contain only one set of reads from either plate, where each set of “Site 1” is labelled identically.
(both called “Site 1” in either manifest, and not “Site 1” and “Site1b” in a single manifest)
You then run the paired end joining as well as the denoising/ debluring steps SEPARATELY, one using the manifest of one run, the second with the other.
THEN, you can combine the table outputs with the above command, and continue with your analysis.
Is this interpretation correct? Or is there a way to combine two samples with unique ID’s after they have been placed in the same table/ rep-seq files using one manifest?
Thank you kindly,
I greatly appreciate the use of your software, as well as your well curated tutorials and forums.